Isolation of a Ca2+-releasing factor from caffeine-treated skeletal muscle fibres and its effect on Ca2+ release from sarcoplasmic reticulum

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Abstract
In the presence of 2mm caffeine, skeletal muscles of the frog exert small irregular oscillations of single sarcomeres. A factor, released from these oscillating muscles, was partially purified, and its activity was tested on skinned fibres and isolated vesicles of the sarcoplasmic reticulum (SR). Purification was achieved in three steps by gel filtration and reversed phase chromatography, and the active compound of the released material was shown probably to be a small peptide. In skinned fibres, the purified factor evoked repetitive contractions and subthreshold sarcomeric oscillations. In ‘heavy’ SR vesicles passively loaded with45Ca2+, the factor induced a small but significant increase in the45Ca2+ efflux rate. At the single channel level, the open probability of the SR Ca2+ release channel increased when the factor was added to the cytoplasmic side of the channel. The results reveal that the released factor potentiates Ca2+ release from the SR by increasing the open time of the Ca2+ release channel.